Technical Field
The present disclosure relates to an apparatus for actuating in rotation and reading a centrifugal microfluidic disk in order to carry out biological and/or biochemical analyses, and to the use of the apparatus for implementing a fluidic protocol.
The present disclosure applies to the field of molecular diagnostics, in particular to LabDisk technology of a monolithic or hybrid type, automating functions such as DNA sequencing, purification of nucleic acids, real-time PCR, and DNA micro-arrays for providing in a single solution automatic systems for analysis of nucleic acids (also known as “sample-in answer-out systems”).
Description of the Related Art
The centrifugal approach to microfluidics offers a unique mode of integrating handling of the liquids for preparing specimens and the subsequent steps of reaction and detection. The process of integration on a single substrate eliminates the need for a separate and preventive handling of the liquids, using pipettes. The characteristics of centrifugal microfluidic systems are particularly attractive in the sectors of life sciences or in applications of in vitro diagnostics where the volumes to be processed in a same test, for example regarding buffer solutions and specimens, frequently differ from one another by several orders of magnitude, and the properties of the various liquids are at times unknown or markedly divergent when specimens different from one another are considered.
However, the above platform has so far encountered little commercial success, as evidenced in the paper by Gorkin R et al., “Centrifugal Microfluidics for Biomedical Applications”, Lab on a Chip, 2010, which presents a treatment of centrifugal microfluidic platforms, highlighting the recent progress in the field and possible future applications.
The major features of disk technology of a hybrid or monolithic type are described in what follows. It is first of all a closed system, presenting ports for inlet and outlet of the specimens, and possibly ventilation ports. Thin disks for containing the liquids are used in order to guarantee a good transfer of heat between the chamber in which the disk is housed and the liquids housed in portions of the disk. The liquids are conveyed into selective locations of the disk by a centrifugal force applied to the disk itself. The processes of cellular lysis, DNA purification, real-time PCR, and other biological reactions take place in wells or chambers of plastic material.
Available in the prior art are numerous disks to be used for centrifugal microfluidics in biological applications. See, for example, the paper by Oliver Strohmeier et al., “Real-Time PCR Based Food Pathogen Detection on a Centrifugal Microfluidic Foil Disk Including Positive- and No-Template-Controls”, 15th International Conference on Miniaturized Systems for Chemistry and Life Sciences, Oct. 2-6, 2011, Seattle, Wash., USA, pp. 506-508.
See, likewise, the paper by M. Focke et al., “Centrifugo-Thermopneumatic Liquid Actuation for Microfluidic Genotyping of Nucleic Acids”, 15th International Conference on Miniaturized Systems for Chemistry and Life Sciences, Oct. 2-6, 2011, Seattle, Wash., USA, pp. 659-661.
See, likewise, the paper by Sascha Lutz et al., “Microfluidic Lab-On-A-Foil for Nucleic Acid Analysis Based on Isothermal Recombinase Polymerase Amplification (RPA)”, Lab on a Chip, 2010, vol. 10, pp. 887-893.
Furthermore, known in the literature are the so-called “miniature stick-packs”, which have a tubular shape and contain reagents in liquid or dry form, to be used in microfluidic disks of a known type. These packs are configured in such a way that they open, releasing the reagents into a reaction chamber of the microfluidic disk, at a precise speed of rotation, or else at a predefined pressure exerted by the reagents that they contain on a seal of the packs themselves. For this purpose, see, for example, the paper by Thomas van Oordt et al., “Miniature Stick-Packaging—An Industrial Technology for Pre-Storage and Release of Reagents in Lab-On-A-Chip Systems”, 15th International Conference on Miniaturized Systems for Chemistry and Life Sciences, Oct. 2-6, 2011, Seattle, Wash., USA, pp. 437-439.
As regards further information on microfluidic disks, and an indication on a method for their manufacture, see, for example, the paper by Maximilian Focke et al., “Microstructuring of Polymer Films for Genotyping by Real-Time PCR on a Centrifugal Microfluidic Platform”, Lab on a Chip, 2010, vol. 10, pp. 2519-2526.
However, the instruments used for carrying out the biological processes using the aforesaid disks show some deficiencies. In particular, the control of temperature within the chamber in which the reactions occur is not optimal in terms of regulation of the temperature and ascending and descending ramps (for example, 0.5° C./s), and the read optics does not enable an adequate sensitivity and spatial resolution to be achieved.